Developing New Tools for Detecting Germline Mutation Induction in Mice

Abstract:

Identifying factors that influence germline mutation rate in animals is an
important toxicological consideration. However, research in this area is limited by the
lack of efficient and precise tools for characterizing germ cell mutagens in humans and
model species such as mice. Thus, the purpose of my research was to develop new tools
for quantifying germline mutation induction in mice. My first objective was to identify
polymorphic microsatellites as a new tool for detecting changes in germline mutation
frequency. Enrichment and DNA sequencing of microsatellites in closely related inbred
mouse lines was used to identify polymorphic loci. The loci were used to screen family
pedigrees of outbred Swiss-Webster mice in which the sires were irradiated. No mutation
induction was detected using the microsatellites; however, the samples used had
previously shown mutation induction at other tandem repeat loci. Failure to detect
mutation induction using the microsatellites does not rule them out as a useful tool, but
rather raises questions as to why induction was detected at other loci but not at
microsatellites. After negative results with microsatellites, I determined if whole genome
sequencing (WGS) using next-generation sequencing (NGS) technologies would be a
suitable alternative for detecting germline mutation induction. WGS is more relevant than
tandem repeat DNA markers to health because it examines all loci, both neutral and
coding, simultaneously. However, NGS is still limited by sequencing error rates, which
are high in comparison to the intergenerational mutation rate, and the high cost associated
with sequencing multiple large genomes. Therefore, WGS is not yet suitable for largescale
germline mutation studies. In conclusion, WGS will be an optimal tool for germline
mutation studies in the near future when costs and error rates decline. WGS will need to be used to determine if there is a correlation between mutation induction at coding
sequence and at neutral marker loci. If a correlation exists, then germline mutation
studies using neutral markers, such as microsatellites, can continue to serve as primary
tools for detecting germline mutation induction.

Description:

A Thesis Submitted to the Faculty of Graduate Studies and Research In Partial Fulfillment of the Requirements for the Degree of Master of Science in Biology, University of Regina. xi, 107 l.